Zhong Li, Wang Yingqun, Kannan Perry, Tainsky Michael A
Program in Molecular Biology and Genetics, Barbara Ann Karmanos Cancer Institute, Wayne State University School of Medicine, 110 East Warren Avenue, Detroit, MI 48201, USA.
Gene. 2003 Nov 27;320:155-64. doi: 10.1016/s0378-1119(03)00823-0.
The transcriptional positive cofactor 4 (PC4) physically interacts with the transcription factor, activator protein-2 (AP-2) alpha, and overexpression of PC4 results in a relief of the AP-2 transcriptional self-interference, which is induced by high levels of AP-2alpha expression. PC4 was initially described as a DNA-binding protein that enhances the activator-dependent transcription of class II genes in vitro, but it was later shown that PC4 could also act as a potent repressor of transcription on specific DNA structures such as single-stranded (ss) DNA, DNA ends and heteroduplex DNA. To further explore the functional domains of PC4 and its ssDNA-binding effect in the interaction with AP-2alpha and on AP-2 transcriptional activity, we investigated the C-terminal domain of PC4 (PC4-CTD) and several PC4 mutants in which the ssDNA binding function was interrupted. We found that the C-terminal domain of PC4 physically interacts with AP-2alpha and retains the function of full-length protein in relieving transcription self-interference of AP-2. A point-mutated form of PC4 within the C-terminal domain beta-ridge, PC4 W89A, or a triple mutant in the beta2-beta3 loop of PC4, F77A/K78G/K80G, inactivate the ability of PC4 to bind AP-2alpha and to relieve the transcription self-interference of AP-2alpha. In addition, point-mutated forms of AP-2alpha within the activation domain (AD) that inactivate AP-2 transcription activity also lose their self-interference function. Our data suggest that the C-terminal domain of the transcription cofactor PC4 is critical for AP-2alpha transcriptional interference that is mediated by the activation domain of AP-2alpha.
转录正向辅因子4（PC4）与转录因子激活蛋白-2（AP-2）α发生物理相互作用，PC4的过表达可缓解由高水平AP-2α表达诱导的AP-2转录自干扰。PC4最初被描述为一种DNA结合蛋白，在体外可增强II类基因的激活因子依赖性转录，但后来发现PC4也可作为特定DNA结构（如单链（ss）DNA、DNA末端和异源双链DNA）转录的有效阻遏物。为了进一步探索PC4的功能结构域及其在与AP-2α相互作用以及对AP-2转录活性中的ssDNA结合效应，我们研究了PC4的C末端结构域（PC4-CTD）以及几个ssDNA结合功能被中断的PC4突变体。我们发现PC4的C末端结构域与AP-2α发生物理相互作用，并保留了全长蛋白缓解AP-2转录自干扰的功能。PC4在C末端结构域β-脊内的点突变形式PC4 W89A，或PC4在β2-β3环中的三突变体F77A/K78G/K80G，使PC4结合AP-2α以及缓解AP-2α转录自干扰的能力失活。此外，激活域（AD）内使AP-2转录活性失活的AP-2α点突变形式也失去了其自干扰功能。我们的数据表明，转录辅因子PC4的C末端结构域对于由AP-2α激活域介导的AP-2α转录干扰至关重要。
